title: Characterisation, immunolocalisation and antifungal activity of a lipid transfer protein from chili pepper (Capsicum annuum) seeds with novel α-amylase inhibitory properties.
abstract: Lipid transfer proteins (LTPs) were thus named because they facilitate the transfer of lipids between membranes in vitro. This study was triggered by the characterization of a 9-kDa LTP from Capsicum annuum seeds that we call Ca-LTP(1) . Ca-LTP(1) was repurified, and in the last chromatographic purification step, propanol was used as the solvent in place of acetonitrile to maintain the protein's biological activity. Bidimensional electrophoresis of the 9-kDa band, which corresponds to the purified Ca-LTP(1) , showed the presence of three isoforms with isoelectric points (pIs) of 6.0, 8.5 and 9.5. Circular dichroism (CD) analysis suggested a predominance of α-helices, as expected for the structure of an LTP family member. LTPs immunorelated to Ca-LTP(1) from C. annuum were also detected by western blotting in exudates released from C. annuum seeds and also in other Capsicum species. The tissue and subcellular localization of Ca-LTP(1) indicated that it was mainly localized within dense vesicles. In addition, isolated Ca-LTP(1) exhibited antifungal activity against Colletotrichum lindemunthianum, and especially against Candida tropicalis, causing several morphological changes to the cells including the formation of pseudohyphae. Ca-LTP(1) also caused the yeast plasma membrane to be permeable to the dye SYTOX green, as verified by fluorescence microscopy. We also found that Ca-LTP(1) is able to inhibit mammalian α-amylase activity in vitro.
authors.affiliation: Laboratório de Fisiologia e Bioquímica de Microorganismos, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense, Campos dos Goytacazes, Rio de Janeiro, Brasil.
authors.forename: Mariângela S
authors.initials: MS
authors.lastname: Diz
authors.forename: Andre O
authors.initials: AO
authors.lastname: Carvalho
authors.forename: Suzanna F F
authors.initials: SF
authors.lastname: Ribeiro
authors.forename: Maura
authors.initials: M
authors.lastname: Da Cunha
authors.forename: Leila
authors.initials: L
authors.lastname: Beltramini
authors.forename: Rosana
authors.initials: R
authors.lastname: Rodrigues
authors.forename: Viviane V
authors.initials: VV
authors.lastname: Nascimento
authors.forename: Olga L T
authors.initials: OL
authors.lastname: Machado
authors.forename: Valdirene M
authors.initials: VM
authors.lastname: Gomes
mesh_terms: D000935:Antifungal Agents / Q000494:pharmacology*; D002212:Capsicum / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology* / Q000648:ultrastructure; D002463:Cell Membrane Permeability / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D056148:Chromatography, Reverse-Phase; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D005658:Fungi / Q000187:drug effects / Q000254:growth & development; D006801:Humans; D008826:Microbial Sensitivity Tests; D010940:Plant Proteins / Q000378:metabolism / Q000648:ultrastructure; D021381:Protein Transport / Q000187:drug effects; D012639:Seeds / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D013045:Species Specificity; D013194:Staining and Labeling; D000516:alpha-Amylases / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D000935:Antifungal Agents; D002352:Carrier Proteins; D010940:Plant Proteins; C030363:lipid transfer protein; D000516:alpha-Amylases